Optimizing E. coli-Based Membrane Protein Production Using Lemo21(DE3) or pReX and GFP-Fusions

  • Grietje Kuipers
  • Markus Peschke
  • Nurzian Bernsel Ismail
  • Anna Hjelm
  • Susan Schlegel
  • David Vikström
  • Joen Luirink
  • Jan-Willem de Gier
Protocol
Part of the Methods in Molecular Biology book series (MIMB, volume 1586)

Abstract

Optimizing the conditions for the production of membrane proteins in E. coli is usually a laborious and time-consuming process. Combining the Lemo21(DE3) strain or the pReX T7-based expression vector with membrane proteins C-terminally fused to Green Fluorescent Protein (GFP) greatly facilitates the optimization of membrane protein production yields. Both Lemo21(DE3) and pReX allow precise regulation of expression intensities of genes encoding membrane proteins, which is critical to identify the optimal production condition for a membrane protein. The use of GFP-fusions allows direct monitoring and visualization of membrane proteins at any stage during the production optimization process.

Key words

Membrane protein Production E. coli Lemo21(DE3) pReX Fluorescence GFP 

Notes

Acknowledgments

This work was supported by grants from the Swedish Research Council, the Carl Tryggers Stiftelse, the Marianne and Marcus Wallenberg Foundation, NIH grant 5R01GM081827-03, and the SSF supported Center for Biomembrane Research to JWdG and the People Programme (Marie Curie Actions) of the European Union’s Seventh Framework Programme FP7/2007-2013/under REA grant agreement n°607072 (NK and MK) and a SystemsX Transition Postdoc Fellowship (SusS).

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Copyright information

© Springer Science+Business Media LLC 2017

Authors and Affiliations

  • Grietje Kuipers
    • 1
    • 2
  • Markus Peschke
    • 3
  • Nurzian Bernsel Ismail
    • 2
  • Anna Hjelm
    • 1
  • Susan Schlegel
    • 4
  • David Vikström
    • 2
  • Joen Luirink
    • 3
  • Jan-Willem de Gier
    • 1
    • 2
  1. 1.Department of Biochemistry and Biophysics, Center for Biomembrane ResearchStockholm UniversityStockholmSweden
  2. 2.Xbrane Biopharma ABSolnaSweden
  3. 3.The Amsterdam Institute of Molecules, Medicines and SystemsVU University AmsterdamAmsterdamThe Netherlands
  4. 4.Molecular Microbial Ecology, Institute of Biogeochemistry and Pollutant DynamicsETH ZurichDübendorfSwitzerland

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